This project has as its general goal the study of cell motility and chemotaxis in cultured cells in vitro in response to exogenous growth factors such as EGF. The results during the previous funding period implicate cofilin in controlling sites of actin polymerization and protrusion in cultured cells. This places cofilin in the unexpected role of a decision maker during chemotaxis helping to determine cell direction in response to chemoattractant. In addition, these results implicate N-WASP as a key regulator of chemotaxis and suggest that cofilin and N-WASP cooperate to regulate cell direction and that both are acting during the earliest barbed end transient in response to EGF. These observations suggest that N-WASP might contribute to chemotaxis through specialized structures at the leading edge that affect adhesion. To evaluate this idea, much more work needs to be done to define the N-WASP dependent actin filament compartment at the leading edge of lamellipods, and its relationship, in space and time, to cofilin activity, upstream regulators of N-WASP, and focal complex formation. The challenge for the future is to understand how cofilin is regulated with high spatial and temporal precision as part of the steering wheel of the cell and how the cofilin regulatory cycle interacts with N-WASP to cause directed cell movement and chemotaxis in culture.